Despite efforts in osteosarcoma (OS) research, the role of inductive moderate hyperthermia (IMH) in delivering and enhancing the antitumor effect of liposomal doxorubicin formulations (LDOX) remains unresolved. This study investigated the effect of a combination treatment with LDOX and IMH on Saos-2 human OS cells. We compared cell viability using a trypan blue assay, apoptosis and reactive oxygen species (ROS) measured by flow cytometry and pro-apoptotic Bax protein expression examined by immunocytochemistry in response to IMH (42 MHz frequency, 15 W power for 30 min), LDOX (0.4 μg/mL), and LDOX plus IMH. The lower IC₅₀ value of LDOX at 72 h indicated increased accumulation of the drug in the OS cells. LDOX plus IMH resulted in a 61% lower cell viability compared to no treatment. Moreover, IMH potentiated the LDOX action on the Saos-2 cells by promoting ROS production at temperatures of <42 °C. There was a 12% increase in cell populations undergoing early apoptosis with a less heterogeneous distribution of Bax after combination treatment compared to those treated with LDOX (p < 0.05). Therefore, we determined that IMH could enhance LDOX delivery and its antitumor effect via altered membrane permeabilization, ROS generation, and a lower level of visualized Bax heterogeneity in the Saos-2 cells, suggesting the potential translation of these findings into in vivo studies.

We compare the effects of an extremely low-frequency electromagnetic field (EMF) with the chemotherapeutic agent doxorubicin (DOX) on tumor growth and the hepatic redox state in Walker-256 carcinosarcoma-bearing rats. Animals were divided into five groups with one control (no tumor) and four tumor-bearing groups: no treatment, DOX, DOX combined with EMF and EMF. While DOX and DOX + EMF provided greater inhibition of tumor growth, treatment with EMF alone resulted in some level of antitumor effect (p < .05). Superoxide dismutase, catalase activity and glutathione content were significantly decreased in the liver of tumor-bearing animals as compared with the control group (p < .05). The decreases in antioxidant defenses accompanied histological findings of suspected liver damage. However, hepatic levels of thiobarbituric acid reactive substances, an indicator of lipid peroxidation, were three times lower in EMF and DOX + EMF groups than in no treatment and DOX (p < .05). EMF and DOX + EMF showed significantly lower activity of serum ALT than DOX alone (p < .05). These results indicate that EMF treatment can inhibit tumor growth, causing less pronounced oxidative stress damage to the liver. Therefore, EMF can be used as a therapeutic strategy to influence the hepatic redox state and combat cancer with reduced side-effects.

Magnetic nanocomposites (MNC) are promising theranostic platforms with tunable physicochemical properties allowing for remote drug delivery and multimodal imaging. Here, we developed doxorubicin-loaded Fe₃O₄–Au MNC (DOX-MNC) using electron beam physical vapor deposition (EB-PVD) in combination with magneto-mechanochemical synthesis to assess their antitumor effect on Walker-256 carcinosarcoma under the influence of a constant magnetic (CMF) and electromagnetic field (EMF) by comparing tumor growth kinetics, magnetic resonance imaging (MRI) scans and electron spin resonance (ESR) spectra. Transmission (TEM) and scanning electron microscopy (SEM) confirmed the formation of spherical magnetite nanoparticles with a discontinuous gold coating that did not significantly affect the ferromagnetic properties of MNC, as measured by vibrating-sample magnetometry (VSM). Tumor-bearing animals were divided into the control (no treatment), conventional doxorubicin (DOX), DOX-MNC and DOX-MNC + CMF + EMF groups. DOX-MNC + CMF + EMF resulted in 14% and 16% inhibition of tumor growth kinetics as compared with DOX and DOX-MNC, respectively. MRI visualization showed more substantial tumor necrotic changes after the combined treatment. Quantitative analysis of T₂-weighted (T₂W) images revealed the lowest value of skewness and a significant increase in tumor intensity in response to DOX-MNC + CMF + EMF as compared with the control (1.4 times), DOX (1.6 times) and DOX-MNC (1.8 times) groups. In addition, the lowest level of nitric oxide determined by ESR was found in DOX-MNC + CMF + EMF tumors, which was close to that of the muscle tissue in the contralateral limb. We propose that the reason for the relationship between the observed changes in MRI and ESR is the hyperfine interaction of nuclear and electron spins in mitochondria, as a source of free radical production. Therefore, these results point to the use of EB-PVD and magneto-mechanochemically synthesized Fe₃O₄–Au MNC loaded with DOX as a potential candidate for cancer magnetic nanotheranostic applications.

Evaluating intratumor heterogeneity with image texture analysis offers a more sophisticated understanding of sarcoma response to treatment. We examined the effects of inductive moderate hyperthermia (IMH) on sarcoma-45 growth and intratumor heterogeneity across tissue, cellular and molecular levels using magnetic resonance imaging (MRI), ultrasound and histology image analysis. IMH (42 MHz, 20 W) inhibited sarcoma-45 growth kinetics by 34% compared to the untreated control group. T₂-weighted MRI brightness was increased by 42%, reflecting more extensive tumor necrosis, while Young’s modulus increased by 37% due to more pronounced connective tissue replacement in response to IMH. Whereas calculations of Moran’s spatial autocorrelation index revealed distinctions in heterogeneity between tumor core, periphery and capsule regions of interest (ROIs) on MRI, ultrasound and histological examination in the untreated tumor-bearing animals, there was no significant difference between core and periphery after IMH. Exposure to IMH increased overall tumor ROI heterogeneity by 22% on MRI but reduced heterogeneity in the core and periphery on ultrasound and histology images. Ki-67 protein distribution was 25% less heterogeneous on the tumor periphery after IMH. Therefore, this study provides a quantitative characterization of IMH effects on different manifestations of intratumor sarcoma-45 heterogeneity using experimental imaging data.